ALS Causative Mutations in KIF5A Disrupt Autoinhibition Leading to Toxic Gain of Function (P8-8.002)

Abstract

<h3>Objective:</h3> The objective of this study was to elucidate the pathogenic mechanisms of ALS causative mutations in motor protein KIF5A. <h3>Background:</h3> KIF5A is a neuronal specific subunit of Kinesin-1, which is a microtubule motor protein that plays roles in axonal transport and cytoskeletal regulation. Mutations in distinct regions of Kinesin-1 lead to a broad range of neurologic diseases. KIF5A enables Kinesin-1 to walk along microtubules and transport various cargos such as mitochondria and RNA granules into the distal axon. ALS-causative mutations in KIF5A affect the structure of the domain of the protein that functions in cargo binding and autoregulation. However, the consequences of ALS-causative mutations on disease pathogenesis are not fully understood. Our central hypothesis was that KIF5A ALS mutations disrupt cargo binding and/or autoregulation leading to neurodegeneration. <h3>Design/Methods:</h3> We generated <i>in vitro</i> models of KIF5A ALS using transient transfection of disease causative variants into neuroblastoma cell lines. We performed immunofluorescence staining and confocal imaging to visualize the distribution of motor proteins KIF5A, KIF5B, and Dynein. We utilized Drosophila S2 cell lines to study mitochondrial motility using live-imaging. <h3>Results:</h3> Whereas wild type (WT) KIF5A displayed relatively homogeneous distribution, ALS mutant KIF5A developed dramatic accumulation within distal neurites. Co-expression of the ALS mutant protein with the WT caused it to accumulate as well. Mitochondria displayed similar mislocalization to distal neurites. No change was seen in the distribution of KIF5B or Dynein. <h3>Conclusions:</h3> We found that ALS mutant KIF5A caused dramatic accumulation of mutant and WT protein as well as mitochondrial within distal neurite tips. These findings suggest that ALS mutation disrupts its autoinhibition causing mislocalization of its cargos through gain of function. These changes in distribution are specific to KIF5A containing motors. Our findings establish dysregulation of KIF5A activity as the underlying pathogenic mechanism for KIF5A ALS. <b>Disclosure:</b> Dr. Brent has received research support from The American Academy of Neurology. Mr. Sterling-Angus has nothing to disclose. Dr. Deng has nothing to disclose.